Degranulation In RBL-2H3 Cells Research Articles

AYA22A Aptamers Mitigate Peanut Allergenicity: Insights from Degranulation Assays and Modulating Immune Responses

Food allergy, particularly peanut allergy (PA), is a growing health concern affecting millions globally. PA can lead to severe reactions, including fatal anaphylaxis. Despite the availability of FDA-approved therapies like Palforzia, a cure remains elusive. Current immunotherapies show promise but lack a definitive cure. This study applies an established computational biology tool to design aptamers targeting Ara h1 and Ara h2. The in silico design aims to streamline the selection process, enabling cost-effective and rapid identification of aptamer candidates. The developed aptamers (AYA22A, including AYA22AR321, AYA22AR211, and AYA22AR524), demonstrated efficacy in inhibiting degranulation of RBL-2H3 cells (rat basophilic leukemia cell line) in vitro. They showed promise in neutralizing peanut allergen-induced immune responses. The selected aptamers inhibited degranulation in RBL-2H3 cells, addressing concerns in raw peanuts. Moreover, these aptamers demonstrated stability and effectiveness in peanut plant seeds and commercial products. Our aptamers exhibited potential in modulating immune responses associated with peanut allergy. They influenced Th1/Th2 balance, indicating a role in cytokine regulation. In vitro studies also showed the aptamers’ impact on immune cell expression and cytokine production, resembling responses observed with established immunotherapies. The findings suggest AYA22A aptamers as a potential therapeutic option for peanut allergy, providing a basis for further in vivo investigations.

The peels of sacred Lotus (Nelumbo nucifera) rhizomes suppress allergic reactions by inhibiting the A23187-induced degranulation in rat basophilic leukemia (RBL[sbnd]2H3) cells

We isolated ten compounds from methanolic extract of the peels of sacred lotus (Nelumbo nucifera) rhizomes which were identified as β-sitosterol linoleate 1, β-sitosterol 2, lupeol 3, stigmasterol 3-O-β-D-glucoside 4, oleanolic acid 5, betulinic acid 6, pinoresinol 7, 4-hydroxybenzoic acid 8, catechin 9 and gallocatechin 10. All of the isolated compounds from the peels of sacred lotus rhizomes are reported for the first time, and were investigated for their anti-allergic activity. We found that three of them, stigmasterol 3-O-β-D-glucoside 4, oleanolic acid 5 and pinoresinol 7, were capable of inhibiting A23187-induced degranulation in RBL-2H3 cells with IC50 values 0.18 ± 0.01 mM, 0.28 ± 0.06 mM, and 0.27 ± 0.01 mM, respectively. With an exception to 4, compounds 5 and 7 achieved the anti-allergic effect without affecting the cells viability even at higher concentrations with their selectivity indices (SI) being >5. By reducing A23187-induced degranulation, it is suggestive of a mechanism attenuation of Ca2+ elevation. Our findings suggest that, the peels of sacred lotus rhizomes would be beneficial for providing an inexpensive source for the production of bioactive compounds with anti-allergic effect.

Shiikuwasha leaf and peel extracts inhibit allergic reactions by suppressing degranulation in RBL-2H3 rat basophilic leukemia cells and immunoglobulin production in mouse spleen lymphocytes.

This study aims to investigate the antiallergic effects of Shiikuwasha (Citrus depressa Hayata) leaf and peel extracts by examining the regulation of degranulation and inflammatory cytokine production from rat basophilic leukemia (RBL-2H3) cells and antigen-specific antibody production in sensitized mouse spleen lymphocytes. In vivo antiallergic activity was evaluated using the passive cutaneous anaphylaxis (PCA) reaction model. Extracts of Shiikuwasha leaves and peel were prepared using 80% methanol and dissolved in dimethylsulfoxide. The dinitrophenyl-human serum albumin-induced β-hexosaminidase levels in immunoglobulin (Ig) E-sensitized RBL-2H3 cells were assessed using enzymatic assays. Cytokine production was measured by enzyme-linked immunosorbent assay. Antibody production capacity was evaluated using lymphocytes isolated from spleens of type I allergy model mice. Lymphocytes were cultured for 72 h with Shiikuwasha extracts, and ovalbumin-specific IgE, IgG1, and IgG2a levels were measured. Shiikuwasha leaf and peel extract significantly reduced β-hexosaminidase release and suppressed interleukin-4 and tumor necrosis factor-α production from RBL-2H3 cells. Ovalbumin-specific IgE and IgG1 production decreased in Shiikuwasha extract-treated lymphocytes. These extracts also significantly suppressed the PCA reaction. Shiikuwasha leaf and peel extract reduce degranulation in RBL-2H3 cells and antibody production in spleen-derived lymphocytes and therefore exhibit antiallergic effects.

Elafibranor PPARα/δ Dual Agonist Ameliorates Ovalbumin-Induced Allergic Asthma.

Asthma is characterized by chronic inflammation and respiratory tract remodeling. Peroxisome proliferator-activated receptors (PPARs) play important roles in the pathogenesis and regulation of chronic inflammatory processes in asthma. The role of PPARγ has been studied using synthetic PPARγ agonists in patients with asthma. However, involvement of PPARα/δ has not been studied in asthma. In the present study, we investigated if elafibranor, a PPARα/δ dual agonist, can modulate ovalbumin (OVA)-induced allergic asthma, which is a potential drug candidate for non-alcoholic fatty liver in obese patients. Elafibranor suppresses antigen-induced degranulation in RBL-2H3 mast cells without inducing cytotoxicity in vitro. In mice with OVA-induced allergic asthma, the administration of elafibranor suppressed OVA-induced airway hyper-responsiveness at a dose of 10 mg/kg. Elafibranor also suppressed the OVA-induced increase in immune cells and pro-inflammatory cytokine production in the bronchoalveolar lavage fluid (BALF). Histological studies suggested that elafibranor suppressed OVA-induced lung inflammation and mucin hyper-production in the bronchial airways. In addition, elafibranor suppressed OVA-induced increases in serum immunoglobulin E and IL-13 levels in BALF. Conversely, the present study suggests that elafibranor has the potential for use in patients with allergic asthma.

Traditional cooking methods decreased the allergenicity of egg proteins.

Egg allergy is one of the most common food allergies globally. This study aimed to assess the impact of four traditional cooking methods on the allergenicity of egg proteins using a comprehensive strategy, including simulated gastrointestinal digestion in vitro, serology experiments, a rat basophilic leukemia (RBL)-2H3 cell degranulation model, and a passive cutaneous anaphylaxis (PCA) mice model, and the structure changes were detected by circular dichroism (CD)spectra and ultraviolet (UV) spectra. The results showed that the processed egg proteins were more readily digested compared to raw egg proteins. The serological experiments revealed a significant reduction in immunoglobulin E binding of egg proteins after thermal treatments (p<0.05), particularly after frying. Subsequently, the RBL-2H3 cell degranulation experiment demonstrated a marked decrease in the level of egg allergens-induced β-hexosaminidase release after cooking (p<0.05). Moreover, the results from the PCA mice model indicated that the increase in vascular permeability was effectively relieved in the treated groups, especially in frying group (p<0.05). Additionally, the α-helix and β-turn contents of processed egg proteins were significantly decreased (p<0.05) compared with native egg proteins. The UV spectra findings showed that all cooking treatments caused significant alterations in the tertiary structure, and fluorescence analysis indicated that cooking decreased the surface hydrophobicity of egg proteins. In conclusion, four traditional cooking methods reduced the allergenicity of egg proteins, particularly frying, and this reduction was associated with structural changes that could contribute to the destruction or masking of epitopes of egg allergens. PRACTICAL APPLICATION: Egg allergy has a serious impact on public health, and there is no ideal treatment method at present. This study demonstrated that four traditional cooking methods (boiling, steaming, baking, and frying) reduced the allergenicity of egg proteins, especially frying, and the results will provide a basis for the development of hypoallergenic egg products.

Degranulation of RBL-2H3 rat basophilic leukemia cells is synergistically inhibited by combined treatment with nobiletin and lactoferrin.

The aim of this study was to elucidate the anti-allergic effects of polymethoxyflavonoids in combination with milk proteins and the mechanism of inhibition. Three polymethoxyflavonoids and two milk proteins were exposed to the rat basophilic leukemia cell line RBL-2H3. β-hexosaminidase was used as an indicator of degranulation inhibition. The mechanism of inhibition was examined by measuring intracellular Ca2+ levels and western blot method. In the degranulation inhibition test with polymethoxyflavonoids and milk proteins alone, nobiletin was the strongest inhibitor in the polymethoxyflavonoid group and lactoferrin in the milk protein group. Next, co-stimulation with nobiletin and lactoferrin showed stronger synergistic degranulation inhibition than treatment with nobiletin or lactoferrin alone. Western blot analysis showed that co-stimulation with nobiletin and lactoferrin significantly downregulated the induction of phospholipase Cγ 1 phosphorylation. The degranulation response in RBL-2H3 cells was synergistically suppressed by co-stimulation of nobiletin and lactoferrin acting on both Ca2+-dependent and Ca2+-independent pathways.

Discovery of pharmacological effects and targets of Citri Grandis Exocarpium based on SYSTCM and virtual screening.

Citri Grandis Exocarpium (Huajuhong, CGE) is the peel of the unripe fruits of Citrus grandis 'Tomentosa' and Citrus grandis (L.) Osbeck, which is commonly used in the clinic for the treatment of cough and indigestion. The pharmacological mechanism of CGE is unclear. In this study, the pharmacological effect of CGE was predicted by System Traditional Chinese Medicine (SYSTCM), which integrated the pharmacological effect prediction approach by artificial intelligence into the systemic traditional Chinese medicine (TCM) platform. The main pharmacological effect of CGE was antiallergy, promoting bile, blood lipid regulation, cardiotonics, diuresis, and antiarrhythmia by prediction of SYSTCM. In vitro cell experiments were carried out to identify the antiallergic effect of CGE. Extracts of Citri Grandis Exocarpium (ECGE) inhibited lipopolysaccharide-induced cell injury and nitric oxide release in RAW264.7 cells. ECGE and naringin-inhibited immunoglobulin E-induced cell degranulation in RBL-2H3 cells. Target profiling, protein interaction network, and molecular docking of compounds from CGE indicated that mitogen-activated protein kinase 14 (MAPK14) and matrix metalloprotease 9 (MMP9) were key potential targets of CGE with antiallergic activity. This study identified and validated the antiallergic effect of CGE by combining SYSTCM, cell experiments, and virtual screening, which provided a new paradigm and approach for studying the pharmacological effect and mechanism of TCM.

Anti-allergic effect of Cyclopia (honeybush) extracts via anti-degranulation activity in a murine allergy model for inhaled antigen.

The anti-allergic effects of extracts prepared from two species of honeybush, Cyclopia genistoides and Cyclopia subternata, were demonstrated in vivo in a murine allergy model for inhaled antigen induced with ovalbumin (OVA) inhalation to mimic pollen allergy. Intake of the extracts increased the production of OVA-specific immunoglobulin (Ig) E (IgE), IgG1, and IgG2a antibodies in serum and significantly suppressed anaphylactic reaction-induced body temperature decline. Moreover, the extracts significantly inhibited antigen-antibody-induced degranulation in RBL-2H3 cells. They also inhibited body temperature decline when the allergic mice were given them after antigen sensitization, indicating that anti-degranulation activity is the major mechanism underlying the anti-allergic effect of Cyclopia extracts. Despite their qualitative and quantitative differences in phenolic composition, the two extracts exhibited similar effects, suggesting that several active compounds might be involved in the activity. Therefore, oral administration of either Cyclopia extract potentially exerts a systemic anti-allergic effect, supporting the increased consumption of honeybush tea for general wellness and improved quality of life.

Formononetin inhibits IgE by huPlasma/PBMCs and mast cells/basophil activation via JAK/STAT/PI3-Akt pathways.

Food allergy is a prevalent disease in the U.S., affecting nearly 30 million people. The primary management strategy for this condition is food avoidance, as limited treatment options are available. The elevation of pathologic IgE and over-reactive mast cells/basophils is a central factor in food allergy anaphylaxis. This study aims to comprehensively evaluate the potential therapeutic mechanisms of a small molecule compound called formononetin in regulating IgE and mast cell activation. In this study, we determined the inhibitory effect of formononetin on the production of human IgE from peripheral blood mononuclear cells of food-allergic patients using ELISA. We also measured formononetin's effect on preventing mast cell degranulation in RBL-2H3 and KU812 cells using beta-hexosaminidase assay. To identify potential targets of formononetin in IgE-mediated diseases, mast cell disorders, and food allergies, we utilized computational modeling to analyze mechanistic targets of formononetin from various databases, including SEA, Swiss Target Prediction, PubChem, Gene Cards, and Mala Cards. We generated a KEGG pathway, Gene Ontology, and Compound Target Pathway Disease Network using these targets. Finally, we used qRT-PCR to measure the gene expression of selected targets in KU812 and U266 cell lines. Formononetin significantly decreased IgE production in IgE-producing human myeloma cells and PBMCs from food-allergic patients in a dose-dependent manner without cytotoxicity. Formononetin decreased beta-hexosaminidase release in RBL-2H3 cells and KU812 cells. Formononetin regulates 25 targets in food allergy, 51 in IgE diseases, and 19 in mast cell diseases. KEGG pathway and gene ontology analysis of targets showed that formononetin regulated disease pathways, primary immunodeficiency, Epstein-Barr Virus, and pathways in cancer. The biological processes regulated by formononetin include B cell proliferation, differentiation, immune response, and activation processes. Compound target pathway disease network identified NFKB1, NFKBIA, STAT1, STAT3, CCND1, TP53, TYK2, and CASP8 as the top targets regulated at a high degree by formononetin. TP53, STAT3, PTPRC, IL2, and CD19 were identified as the proteins mostly targeted by formononetin. qPCR validated genes of Formononetin molecular targets of IgE regulation in U266 cells and KU812 cells. In U266 cells, formononetin was found to significantly increase the gene expression of NFKBIA, TP53, and BCL-2 while decreasing the gene expression of BTK TYK, CASP8, STAT3, CCND1, STAT1, NFKB1, IL7R. In basophils KU812 cells, formononetin significantly increased the gene expression of NFKBIA, TP53, and BCL-2 while decreasing the gene expression of BTK, TYK, CASP8, STAT3, CCND1, STAT1, NFKB1, IL7R. These findings comprehensively present formononetin's mechanisms in regulating IgE production in plasma cells and degranulation in mast cells.

Paedoksan ameliorates allergic disease through inhibition of the phosphorylation of STAT6 in DNCB-induced atopic dermatitis like mice

Various allergic diseases such as atopic dermatitis (AD), allergic rhinitis, and asthma are considered incurable conditions that have yet to be fully conquered. Paedoksan (PDS), an herbal preparation consisting of 14 medicines, displays effective anti-inflammatory and anti-allergic properties, yet its underlying molecular mechanism is unknown. This study aims to uncover PDS’s mechanism for treating allergic diseases and suggest its therapeutic potential. Through a network pharmacological prediction, its impact on signal transducer and activator of transcription 6 (STAT6) regulation, a sub-mechanism of interleukin 4 (IL-4), a major inflammatory cytokine involved in degranulation and allergy, was investigated in RBL-2H3 cells and an atopic mouse model. PDS inhibits immunoglobulin E (IgE)-induced degranulation and STAT6 phosphorylation evoked by IL-4 in granulocytes. The downregulation of phospho-STAT6 and thymic stromal lymphopoietin (TSLP) by PDS was confirmed in 2,4-dinitrochlorobenzene (DNCB)-induced mouse skin. The results demonstrate that PDS exhibited remarkable effects on degranulation and STAT6 phosphorylation in RBL-2H3 cells, as well as in an atopic mouse model. Furthermore, the main active components from PDS based on chromatographic analysis showed good accordance with PDS’s effects on RBL-2H3 cells. In summary, these findings collectively suggest that PDS holds the potential to effectively suppress inflammatory and allergic reactions by obstructing the target IL-4 protein and its downstream effects, as elucidated through a network pharmacological analysis.

Effects and potential mechanisms of Saposhnikovia divaricata (Turcz.) Schischk. On type I allergy and pseudoallergic reactions in vitro and in vivo

Ethnopharmacological relevanceThe incidence of allergic disease is constantly increasing, but its pathogenesis is not fully understood. Saposhnikovia divaricata (SD), called ‘Fangfeng’ in China, not only can be used for antipyretic, analgesic and anti-inflammatory as a traditional Chinese medicine, but also as an active ingredient in about 8% prescriptions. However, its effects on type I allergy and pseudoallergy have not been clarified. Aim of the studyTo explore the treatment and potential mechanisms of SD and its major bioactive component Prim-O-glucosylcimifugin (POG) on type I allergy and pseudoallergy in vitro and in vivo. Materials and methodsThe inhibitory effect of SD decoction and POG on type I allergy and its possible mechanism were evaluated by using RBL-2H3 cells model in vitro and the passive cutaneous anaphylaxis (PCA) mouse model in vivo. The cell degranulation of RBL-2H3 cells induced by DNP-IgE/DNP-BSA and Compound 48/80 (C48/80) was investigated, and the molecules of degranulation related signaling pathway was further detected by qRT-PCR and Western Blot analysis. Meanwhile, therapeutic effect of SD Decoction and POG were evaluated using PCA models in vivo. The molecular docking technology was conducted to explore the potential mechanisms. ResultsIn cells model induced by DNP-IgE/DNP-BSA, the release rate of β-Hex in high dose of SD and POG groups were 43.79% and 57.01%, and the release amount of HA in high dose of SD and POG groups were 26.19 ng/mL and 24.20 ng/mL. They were significantly lower than that in the model group. Besides, SD decoction and POG could significantly inhibit intracellular Ca2+ increasing and cell apoptosis. But there is no obvious effect on cells degranulation induced by C48/80. The molecular docking results showed that 5-O-Methylvisamioside and POG could bind with FcεRI α with stronger binding ability, but weak binding ability to Mrgprx2. Moreover, qPCR and Western blot analyses indicated that SD could down-regulate Lyn/Syk/PLCγ, MAPK and PI3K/AKT/NF-κB signal pathway to inhibit IgE-dependent cell degranulation. In mice PCA model, both SD and POG could dose-dependently attenuate the Evans Blue extravasation, paw and ear swelling induced by DNP-IgE/DNP-BSA, but no significant inhibition under the PCA models induced by C48/80. ConclusionIn conclusion, SD is effective for the therapeutic of type I allergies, suggesting that SD is a potential candidate for the treatment of type I allergy, and the underlying mechanism of these effects needs to be further studied.

Isolation and Quantification of 2-O-Caffeoyl-hydroxycitric Acid, an Active Component in Hot Water Extracts of Coriander (Coriandrum sativum L.), Which Inhibits Degranulation of RBL-2H3 Cells

Objectives: Natural compounds that can effectively prevent and alleviate allergic symptoms, such as hay fever, are widely sought after from sources such as natural ingredients that can be ingested daily. Coriander extracts have been reported to show anti-allergic activity; however, hot water extracts have not been tested previously. Here, we found that a hot water extract of the aerial parts of coriander inhibited the degranulation of rat basophilic leukemia cell line RBL-2H3 cells and identified the active compounds. Methods: Chromatographic methods were used to isolate compounds from the hot water extract of coriander. The chemical structures were elucidated by analyses of high-resolution electrospray ionization mass spectrum and nuclear magnetic resonance spectral data. The degranulation activity of compounds was evaluated using the β-hexosaminidase release assay. Results: We analyzed the components to identify active compounds. 2 S,3 S-2- O-caffeoyl-hydroxycitric acid (1), and 2- O-caffeoyl-hydroxycitric acid 6′- O-methyl ester (2) were isolated from the hot water extract of coriander, and both compounds showed degranulation inhibitory activity in RBL-2H3 cells at a concentration below 2.5 mM. The content of compound 1 in the freeze-dried coriander powder was calculated to be 2.0% via HPLC quantification. Conclusions: Compound 1 was considered to be the primary active contributor in the hot water extract of coriander.

Effects of Simvastatin on RBL-2H3 Cell Degranulation.

Hypercholesterolemia is a major complication of arteriosclerosis. Mast cells in arteriosclerosis plaques induce inflammatory reactions and promote arterial sclerosis. In this study, we evaluated the pharmacological effects of simvastatin (SV)-3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase inhibitors on the degranulation of rat basophilic leukemia (RBL)-2H3 cells, which are commonly used as mast cell models. SV significantly decreased the degranulation induced by three types of stimulation: antigen antibody reaction (Ag-Ab), thapsigargin (Tg) serosal endoplasmic reticulum calcium ATPase (SERCA) inhibitor, and A23187 calcium ionophore. SV had a stronger inhibitory effect on degranulation induced by Ag-Ab stimulation than the other two stimulations. However, SV did not inhibit increase of intracellular Ca2+ concentrations. Mevalonate or geranylgeraniol co-treatment with SV completely prevented the inhibitory effect of SV on the degranulation induced by these stimulations. Immunoblotting results showed that SV inhibited protein kinase C (PKC) delta translocation induced by Ag-Ab but not by Tg or A23187. SV induced a reduction in active Rac1, and actin filament rearrangement. In conclusion, SV inhibits RBL-2H3 cell degranulation by inhibiting downstream signaling pathways, including the sequential degranulation pathway. These inhibitory effects were completely reversed by the addition of geranylgeraniol and might be induced by changes in the translocation of the small guanosine 5'-triphosphatase (GTPase) families Rab and Rho, which are related to vesicular transport PKC delta translocation and actin filament formation, respectively. These changes are caused by the inhibition of HMG-CoA reductase by SV following the synthesis of geranylgeranyl pyrophosphates, which play important roles in the activation of small GTPases, Rab.

Assessment of the effect of glycation on the allergenicity of sesame proteins

Sesame allergy is a growing concern worldwide. In this study, sesame proteins was glycated with glucose, galactose, lactose and sucrose respectively, and the allergenicity of different glycated sesame proteins were assessed by a comprehensive strategy, including simulated gastrointestinal digestion in vitro, a BALB/c mice model, a rat basophilic leukemia (RBL)–2H3 cell degranulation model and a serological experiment. Firstly, simulated gastrointestinal digestion in vitro showed that glycated sesame proteins were more easily to digest than raw sesame. Subsequently, the allergenicity of sesame proteins was assessed in vivo by detecting the allergic indexes of mice, and results showed that the levels of total immunoglobulin E (IgE) and histamine were reduced in glycated sesame proteins treated mice. Meanwhile, the Th2 cytokines (IL-4, IL-5, and IL-13) were downregulated significantly, demonstrating that sesame allergy was relieved in glycated sesame treated mice. Thirdly, the RBL-2H3 cell degranulation model results showed that the release of β-hexosaminidase and histamine were decreased to different degrees in glycated sesame proteins treated groups. Notably, the monosaccharide glycated sesame proteins exhibited lower allergenicity both in vivo and in vitro. Furthermore, the study also analyzed the structure alteration of sesame proteins, and the results showed that the secondary structure of glycated sesame proteins were changed (the content of α-helix and β-sheet were reduced), and the tertiary structure of sesame proteins after glycation modification was also changed (microenvironment around aromatic amino acids was altered). Besides, the surface hydrophobicity of glycated sesame proteins was also reduced except sucrose glycated sesame proteins. In conclusion, this study demonstrated that glycation reduced the allergenicity of sesame proteins effectively, especially glycation with monosaccharides, and the allergenicity reduction might be related to structural changes. The results will provide a new reference for developing hypoallergenic sesame products.

Synthesis and structure-activity relationship of kujigamberol B, a dinorlabdane diterpenoid isolated from an ancient Kuji amber.

The versatile methodology was developed for synthesizing kujigamberol B, a dinorlabdane diterpenoid isolated from the methanol extract of Kuji amber. A highly efficient intramolecular cyclization is followed by a Sonogashira-coupling reaction during the total synthesis. The synthesized compounds were evaluated for the growth-restoring activity against the mutant yeast (zds1Δ erg3Δ pdr1Δ pdr3Δ) and for the degranulation of RBL-2H3 cells. We found that in both activities, primary alcohol and secondary alcohol analogs are as active as kujigamberol B.

Effects of Lactobacillus fermentation on Eucheuma spinosum polysaccharides: Characterization and mast cell membrane stabilizing activity

Eucheuma polysaccharides have varieties of biological activities. However, it is accompanied by problems like large molecular weight, high viscosity, and low utilization. Here, we first prepared fermented Eucheuma spinosum polysaccharides (F-ESP) by Lactobacillus fermentation, compared with low-temperature freeze-thaw ESP (L-ESP) prepared by the freeze-thaw method, explored the composition and structural characteristics of F-ESP and L-ESP, and evaluation of the ability of different samples to inhibit mast cell degranulation using classical mast cell model. Then, the activity of L-ESP and F-ESP in vivo was preliminarily evaluated using a passive cutaneous anaphylaxis model. Two kinds of F-ESP named F1-ESP-3 and F2-ESP-3 were obtained by fermentation of Eucheuma spinosum with the selected strains of Lactobacillus.sakei subsp.sakei and Lactobacillus.rhamnosus. Compared with the purified component L-ESP-3, the monosaccharide composition of F1-ESP-3 contains more glucuronic acid, the molecular weight reduced from >600 kDa (L-ESP-3) to 28.30 kDa (F1-ESP-3) and 33.58 kDa (F2-ESP-3), F1-ESP-3 has higher solubility and lower apparent viscosity. Fermentation did not destroy the functional groups and structure of ESP. Moreover, F1-ESP-3 significantly inhibited RBL-2H3 cell degranulation by reducing depolymerization of F-actin and Ca2+ influx. F1-ESP-3 reduced the symptoms of mast cell-mediated passive cutaneous anaphylaxis, indicating that F1-ESP-3 may have better anti-allergic activity in vivo.

Ultramicronized N-Palmitoylethanolamine Regulates Mast Cell-Astrocyte Crosstalk: A New Potential Mechanism Underlying the Inhibition of Morphine Tolerance.

Persistent pain can be managed with opioids, but their use is limited by the onset of tolerance. Ultramicronized N-palmitoylethanolamine (PEA) in vivo delays morphine tolerance with mechanisms that are still unclear. Since glial cells are involved in opioid tolerance and mast cells (MCs) are pivotal targets of PEA, we hypothesized that a potential mechanism by which PEA delays opioid tolerance might depend on the control of the crosstalk between these cells. Morphine treatment (30 μM, 30 min) significantly increased MC degranulation of RBL-2H3 cells, which was prevented by pre-treatment with PEA (100 μM, 18 h), as evaluated by β-hexosaminidase assay and histamine quantification. The impact of RBL-2H3 secretome on glial cells was studied. Six-hour incubation of astrocytes with control RBL-2H3-conditioned medium, and even more so co-incubation with morphine, enhanced CCL2, IL-1β, IL-6, Serpina3n, EAAT2 and GFAP mRNA levels. The response was significantly prevented by the secretome from PEA pre-treated RBL-2H3, except for GFAP, which was further upregulated, suggesting a selective modulation of glial signaling. In conclusion, ultramicronized PEA down-modulated both morphine-induced MC degranulation and the expression of inflammatory and pain-related genes from astrocytes challenged with RBL-2H3 medium, suggesting that PEA may delay morphine tolerance, regulating MC-astrocyte crosstalk.

Sea cucumber chondroitin sulfate polysaccharides attenuate OVA-induced food allergy in BALB/c mice associated with gut microbiota metabolism and Treg cell differentiation.

Previous research studies have shown that sulfated polysaccharides can inhibit food allergy, but the detailed mechanism remains largely unknown. In this study, RBL-2H3 cells were used to compare the anti-allergic activities of four sulfated polysaccharides, and an ovalbumin (OVA)-sensitized allergic mouse experiment was used to explore their desensitization effect, with regard to the alteration in gut microbiota and immune cell differentiation. Compared with the shark, bovine and porcine chondroitin sulfate, sea cucumber chondroitin sulfate (SCCS) significantly inhibited the degranulation of RBL-2H3 cells. SCCS reduced allergic symptoms and protected the jejunum from injury in mice. Furthermore, SCCS increased the relative abundance of Lachnospiraceae NK4A136, decreased the relative proportion of Prevotellaceae NK3B31, and up-regulated the secretion of short chain fatty acids such as butyric acid in the feces, resulting in an increase in the mucin 2 (MUC2) secretion by goblet cells HT-29. Meanwhile, SCCS induced the differentiation of regulatory T cells in the mesenteric lymph nodes of mice. This study provides a deeper understanding of the functioning mechanism of SCCS in alleviating food allergy and may guide the development and production of anti-allergy active ingredients.

The Heterogeneity of Ovomucoid-Specific IgE Idiotype Is Associated With Egg Allergy Symptom Severity.

Immunoglobulin E (IgE)-mediated egg allergy presents as one of the most common food allergies. The level of specific IgE (sIgE) antibody is widely used as an important in vitro diagnostic indicator. However, sIgE antibody levels are often inconsistent with the clinical manifestations of patients. The heterogeneity of egg-specific IgE idiotypes (sIgE-IDs) may help reflect clinical egg allergy severity. Eight peptides were synthesized, corresponding to the linear epitopes of ovomucoid (OVM). The sIgE-IDs of egg-allergic patients were detected by enzyme-linked immunosorbent assay. Fresh peripheral blood was collected from patients with different heterogeneity strength of sIgE-ID, and egg extract was used as a stimulus to the basophil activation test (BAT). RBL-2H3 cells were sensitized with serum with different strength of sIgE-ID heterogeneity and the release rate of β-hexosaminidase was calculated. Among 75 patients with egg allergy, 24% had sIgE for all epitopes and 85% had sIgE for at least one epitope. Analysis of individual patients revealed differences in epitope recognition patterns among the patients, that is, heterogeneity in sIgE-ID. More importantly, the number of IgE-positive peptides had a strong correlation with allergic symptoms in egg-allergic patients (r = 0.706). BAT and RBL-2H3 cell degranulation confirmed that higher sIgE-ID heterogeneity strength was more effective in inducing effector cell responses. Our results suggest that the greater the heterogeneity strength of OVM-sIgE-ID, the more severe the allergic symptoms.

Protection of Allergic Asthma in Mice by Black Rice Bran Bioprocessed with Shiitake Mushroom Mycelia

We evaluated the potential of orally fed new food formulations to inhibit biomarkers reported to be involved in the causes of allergic asthma in mice. Asthma, a serious non-communicable disease, affects both adults and children and can be undertreated. New functional foods could provide therapeutic approaches. Here, the anti-asthma mechanism of a new functional food and three isolated fractions produced by bioprocessing black rice bran with shiitake mushroom mycelia was evaluated in mast cells, B cells, and orally fed mice and compared with non-bioprocessed black rice bran. In vitro, the treatments inhibited RBL-2H3 cell degranulation and immunoglobulin E (IgE) production. The in vitro anti-asthma effects were confirmed in orally fed mice following asthma induction by alumina and chicken egg ovalbumin (OVA). The suppression of asthma resulted from the inhibition of inflammation- and immune-related substances, including OVA-specific IgE, thymic stromal lymphopoietin, eotaxin, leukotriene C4, prostaglandin D2, and vascular cell adhesion molecule-1 in bronchoalveolar lavage fluid and serum. The treatment also reversed the thickening of the lung airway wall. The inflammation and asthma inhibition seems to be regulated by the balance of the T-helper cells’ Th1/Th2 immune response and the inhibition of multiple biomarkers associated with the cause of asthma. Future human clinical studies with adults and children should determine the potential therapeutic value of the anti-asthma effects of the new functional foods.